The long-term goal of our laboratory is to elucidate the role of inositol polyphosphates (InsPs) in transcriptional regulation. InsPs are generated by phospholipase C (PLC) - dependent pathway that starts with PLC-catalyzed hydrolysis of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] to yield inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and diacylglycerol (DAG). Since Plc1p is the only PLC enzyme in the budding yeast Saccharomyces cerevisiae, and hydrolysis of PtdIns(4,5)P2 is the only pathway for synthesis of Ins(1,4,5)P3 and other InsPs (York et al., 1999), cells with deletion of PLC1 gene (plc1) are completely devoid of all InsPs and the budding yeast thus represents an ideal system in which to study metabolism and cellular roles of InsPs. InsPs affect transcriptional control (Odom et al., 2000) and regulate activity of chromatin remodeling complexes (Shen et al., 2003;Steger et al., 2003). Our recent data indicate that plc1 cells display several phenotypes consistent with the role of Plc1p and InsPs in histone acetylation: (i) defect in recruitment of the SAGA complex to Sko1p-regulated promoters (Guha et al., 2007), (ii) decreased histone H3 and H4 acetylation in the promoters of ribosomal protein genes and FLR1 gene, and (iii) reduced cellular level of acetylated histone H3 and H4. Based on these data, we propose to test the hypothesis that Plc1p and InsPs affect histone acetylation. The Specific Aim 1 will test the hypothesis that Plc1p and InsPs are required for the induction of the FLR1 gene by supporting acetylation of histones H3 and H4 and thus facilitating retention of Swi/Snf complex at the FLR1 promoter. The Specific Aim 2 will determine whether Plc1p and InsPs are required for full expression, acetylation of histones H3 and H4 in the promoter regions, and recruitment of SAGA, Swi/Snf, and NuA4 complexes during induction of GAL1 gene and other inducible genes. We will also determine whether recombinant subcomplexes SAGA and NuA4 can bind InsPs and whether InsPs stimulate SAGA- and NuA4- mediated acetylation of histones and/or nucleosome arrays in vitro . The Specific Aim3 will test the hypothesis that Plc1p and InsPs are required for normal level of histone H4 acetylation at lysine 16 (H4K16) that is required to antagonize the spread of silenced chromatin. Histone acetylation affects chromatin structure and regulates diverse cellular functions, such as gene expression, DNA repair, and cell proliferation. Many histone deacetylase inhibitors reactivate the transcription of multiple genes that are silenced in human tumors and thus possess anti-cancer activity (Kim et al., 2006). Elucidation of the role(s) of InsPs in histone acetylation and transcriptional regulation will contribute to the identification of novel molecular targets for cancer therapy. PUBLIC HEALTH RELEVANCE: Balanced growth of cells requires their accurate transcriptional responses to nutritional and other environmental conditions. Many of these responses require phospholipase C (PLC) and errors in these responses often result in cancer. Elucidation of the regulatory interactions between PLC signaling pathway and transcriptional machinery is important for the identification of novel molecular targets for cancer therapy.